Natural and Unanticipated Modifiers of RNAi Activity in Caenorhabditis elegans

Abstract

Organisms used as model genomics systems are maintained as isogenic strains, yet evidence of sequence differences between independently maintained wild-type stocks has been substantiated by whole-genome resequencing data and strain-specific phenotypes. Sequence differences may arise from replication errors, transposon mobilization, meiotic gene conversion, or environmental or chemical assault on the genome. Low frequency alleles or mutations with modest effects on phenotypes can contribute to natural variation, and it has proven possible for such sequences to become fixed by adapted evolutionary enrichment and identified by resequencing. Our objective was to identify and analyze single locus genetic defects leading to RNAi resistance in isogenic strains of Caenorhabditis elegans. In so doing, we uncovered a mutation that arose de novo in an existing strain, which initially frustrated our phenotypic analysis. We also report experimental, environmental, and genetic conditions that can complicate phenotypic analysis of RNAi pathway defects. These observations highlight the potential for unanticipated mutations, coupled with genetic and environmental phenomena, to enhance or suppress the effects of known mutations and cause variation between wild-type strains.

Systematic comparison of bacterial feeding strains for increased yield of Caenorhabditis elegans males by RNA interference‐induced non‐disjunction

“…While the Him phenocopies produced in our experiments were reproducible, the experimental deviations highlight the intrinsically variable nature of RNAi by feeding (Table 2). A number of different factors may contribute to experimental inconsistency: batch-to-batch variability in the accumulation of dsRNA within the bacterial strains, animal-to-animal differences in the amount of dsRNA ingested or the amount that is made bioavailable, or RNAi response differences in individual animals due to unanticipated background mutations in genes with RNAi-related functions [20][21][22][23]. For the particular case of producing Him phenocopies, the presence of males may not directly reflect the effectiveness of the knockdown.…”

“…This list should serve as a good starting point for the production of large numbers of males from different strains. However, lab-to-lab as well as strain-to-strain variability should be anticipated, and investigators should realize the potential of background mutations, including those that affect RNAi [20][21][22][23]. The number of effective ''male foods'' identified improves flexibility in experimental design; for example, selective use of certain ''male foods'' may help avoid interactions between the RNAi phenocopy and mutations in the strain of interest, as well as improve male fertility, as we demonstrated for mut-7 and rde-2 males.…”